title: In vivo analysis of aquaporin 0 function in zebrafish: permeability regulation is required for lens transparency.
abstract: OBJECTIVE
  The zebrafish lens is well suited for studies of physiology and development due to its rapid formation in the embryo and genetic accessibility. Aquaporin 0 (AQP0), a lens-specific membrane protein, is required for lens clarity. Zebrafish have two copies of AQP0 (Aqp0a and b), whereas mammals have a single, multifunctional protein. Here we demonstrate a reliable knockdown/rescue system in zebrafish and use it to provide evidence for subfunctionalization of Aqp0a and b, as well as to show that calcium-mediated regulation of Aqp0a in zebrafish lenses is necessary for transparency.
  
  
  METHODS
  Coinjection of antisense oligonucleotides and DNA rescue constructs into zebrafish embryos, followed by evaluation of the developing fish for cataracts, was used to analyze the functions of Aqp0a and b. The water permeability and regulation characteristics of each rescue protein were tested in a Xenopus oocyte swelling assay.
  
  
  RESULTS
  Both copies of AQP0 are necessary for lens clarity in the zebrafish, and neither is sufficient. Water permeability is necessary but also insufficient. Phosphorylation and regulation of Aqp0a are required for its function.
  
  
  CONCLUSIONS
  In the zebrafish lens, the two closely related AQP0s have acquired distinct functions that are both necessary for lens development and clarity. Regulation of AQP0 water permeability, a well-studied phenomenon in vitro, may be physiologically relevant in the living lens.
authors.affiliation: Department of Physiology and Biophysics, University of California, Irvine, Irvine, California 92697-4561, USA.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Clemens
authors.forename: Karin L
authors.initials: KL
authors.lastname: Németh-Cahalan
authors.forename: Lien
authors.initials: L
authors.lastname: Trinh
authors.forename: Tailin
authors.initials: T
authors.lastname: Zhang
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Schilling
authors.forename: James E
authors.initials: JE
authors.lastname: Hall
mesh_terms: D000818:Animals; D020346:Aquaporins / Q000502:physiology*; D001692:Biological Transport / Q000502:physiology; D002386:Cataract / Q000378:metabolism / Q000503:physiopathology*; D004195:Disease Models, Animal; D005136:Eye Proteins / Q000502:physiology*; D007908:Lens, Crystalline / Q000378:metabolism / Q000502:physiology*; D010539:Permeability; D014867:Water / Q000378:metabolism; D015027:Zebrafish / Q000502:physiology*
chemical_list: D020346:Aquaporins; D005136:Eye Proteins; C030328:aquaporin 0; D014867:Water
keywords: AQP0; cataract; subfunctionalization; zebrafish